新城疫活疫苗HB92克隆株的筛选、纯化与细胞培养特性观察

Selection, Purification and Cell Cultural Characterization of Newcastle Disease Virus(NDV) Live Vaccine Strain HB92

将引进的新城疫V4无毒耐热毒株连续用鸡胚传代,每次挑选48～96 h有明显病变的死亡鸡胚,取其尿囊液毒于-20℃保存,观察V4毒株对鸡胚毒力的病理变化。结果显示,经过20代次的鸡胚毒力筛选,V4毒株对鸡胚的致死率从小于10%上升至20%以上,接种后120 h的活胚有明显地萎缩症状。将鸡胚20代次以上筛选的V4病毒液在鸡胚成纤维细胞上传代培养,并进行蚀斑筛选纯化病毒,直到第5代才出现明显地细胞病变。使用二层琼脂进行病毒的蚀斑分析,第二层琼脂糖在第一层凝固后72 h覆盖,8 h后NDV鸡胚成纤维细胞适应株的蚀斑出现,蚀斑大小直径在1～2 mm变化。通过对NDV V4鸡胚毒力筛选和蚀斑纯化,获得了一株蚀斑克隆株(NDV HB92)。

The avirulent Australia NDV strain V4 with heat-resistant was used in the development of vaccine. To improve its immunogenicity, the pathological change of embryo influenced by NDV was observed. Results showed that after 20-time virulent selections by chick embryo cuhure, the fatality rate of chick embryo increased nearly 10%. When the eggs with NDV were incubated for 120 hours, the atrophy embryo was observed. The virulence-improved NDV V4 was further incubated in chick embryo fibroblast and purified, obviously cytopathic effect appeared in the 5^th passage. Finally one new NDV straiu HB92 was obtained by chick embryo virulent selection and plague purification.